GLUTATHIONE ANALOGS AS NOVEL INHIBITORS OF RAT AND HUMAN GLUTATHIONE-S-TRANSFERASE ISOENZYMES, AS WELL AS OF GLUTATHIONE CONJUGATION IN ISOLATED RAT HEPATOCYTES AND IN THE RAT IN-VIVO

Inhibitors of rat and human Alpha- and Mu-class glutathione S-transfer ases that effectively inhibit the glutathione (GSH) conjugation of bro mosulphophthalein in the rat liver cytosolic fraction, isolated rat he patocytes and in the rat liver in vivo have been developed. The GSH an alogue carboxy-2-gamma-(S)-glutamylamino-N-hexylpentamide [Adang, Brus see, van der Gen and Mulder (1991) J. Biol. Chem. 266, 830-836] was us ed as the lead compound. To obtain more potent inhibitors, it was modi fied by replacement of the N-hexyl moiety by N-2-heptyl and by esterif ication of the 5-carboxy group with ethyl and dodecyl groups. In isola ted hepatocytes, the branched N-2-heptyl derivatives were stronger inh ibitors of GSH conjugation of bromosulphophthalein than the N-hexyl de rivatives. The ethyl ester compounds were more efficient than the corr esponding unesterified derivatives. The dodecyl ester of the N-2-hepty l analogue was the most effective inhibitor in isolated hepatocytes, b ut was relatively toxic in vivo. However, the corresponding ethyl este r was a potent in vivo inhibitor: GSH conjugation of bromosulphophthal ein (as assessed by biliary excretion of the conjugate) was decreased by 70% after administration of a dose of 200 mu mol/kg. The isoenzyme specificity of the inhibitors towards purified rat and human glutathio ne S-transferases was also examined. The unesterified compounds were m ore potent than the esterified analogues, and inhibited Alpha- and Mu- class isoenzymes of both rat and human glutathione S-transferase (K-i range 1-40 mu M). Other GSH-dependent enzymes, i.e, GSH peroxidase, GS H reductase and gamma-glutamyltranspeptide, were not inhibited. Thus l oxycarbonyl-2-gamma-(S)-glutamylamino-N-2-heptyl- pentamide, the in vi vo inhibitor of GSH conjugation, may be useful in helping to assess th e role of the Alpha and Mu classes of glutathione S-transferases in ce llular biochemistry, physiology and pathology.